ABSTRACT Reptiles are a long-lived vertebrate group often overlooked in immunology research. The humoral immune system of reptiles contains both innate and adaptive components, but the organization of those components differs, for example reptiles lack lymph nodes, but have gut-associated lymphoid tissue. In addition, researchers are just beginning to appreciate the nature of the humoral immune response, particularly B cell function in reptiles. Given the many similarities of human and reptilian humoral immune systems, it is surprising then that long-lived reptiles appear to maintain immunocompetence throughout the aging process, unlike humans. Additionally, virtually nothing is known about mucosal humoral immunity in reptiles, and is likely to be critical for preventing infection during old age through production of secreted antibodies. This project expands our previous research on B cell function in the red-eared slider, Trachemys scripta. We and our students have previously found B cells are capable of two distinct functions: production of antibodies (spontaneous natural antibodies and antigen-induced specific antibodies) and phagocytosis. While reptilian B cells appear similar to mammalian B-1 cells, it is currently unclear if separate subsets of cells carry out each function or if a single cell can choose its fate. The proposed research will address two questions: (1) How are reptilian B cells directed toward each of their functions? and (2) What is the distribution and function of reptilian mucosal B cells? Using a natural population of turtles of various ages, we'll address these questions with ELISA, ELISpot, and phagocytic assays to test the responses of B cells at the individual cell and population levels. We will also examine the age-related distribution and function of B cells within the intestinal mucosa, expanding our knowledge of how reptiles use these tissues to defend against pathogens. This project will allow the PIs to continue their excellent record of student training in hands-on research. Undergraduate and graduate students will be involved in all aspects of the project and this experience will provide them opportunities they would not otherwise have to develop successful future research careers. The results of these experiments will provide important insight into the function and distribution of reptilian B cells, and provide novel information on mucosal immunity in this understudied group. Understanding the successful immune strategies of these long-lived animals could provide new interventions to extend human health span.